There is a strong correlation between chronic drug use and increased susceptibility to HIV infection. Chronic drug users account for approximately one third of all cases of HIV in the USA and progression to AIDS is markedly accelerated in opiate drug abusers. Chronic immune activation with increased serum levels of proinflammatory cytokines is a hallmark of progressive disease. Recent studies show that elevated plasma endotoxin (LPS), a consequence of gut bacterial translocation, may be the likely cause of immune activation in HIV infection. A correlation has been found between immune activation and T cell activation/depletion. Interestingly, studies show that HIV patients that use intravenous heroin display higher serum levels of LPS when compared to non-drug using HIV infected patients. Similarly, we show in a murine model of drug abuse, higher circulating levels of endotoxin when compared to placebo treated animals. These effects are further potentiated in the TATtg mice. As observed in human patients, immune activation in these animal models were also prolonged and sustained. These results implicate a role for LPS in driving the disease progression in drug abusing HIV infected patients. Paradoxically, however, it is well documented that, prolonged LPS exposure leads to LPS tolerance with decreased LPS induced macrophage stimulation. Yet, sustained immune activation is a hallmark of disease progression in both HIV patients and in animal models of HIV. The mechanism underlying this discordant observation is a significant gap in knowledge. In LPS tolerance, transcriptional and translational repressive events combine to tightly regulate proinflammatory genes. More recently, microRNAs have been implicated as negative regulators controlling diverse biological processes at the level of post-transcriptional regulation. We show in preliminary data a significant induction of two key miRNAs, miR-155 and miR-146a in animals that are chronically treated with LPS. In contrast, both miR-155 and miR-146a are significantly down regulated in animals that are morphine or TAT treated. LPS activates Toll like receptor 4 to induce transcription of proinflammatory cytokines and chemokines (32-34). LPS induction of miR-155 and miR-146a may act as negative feedback regulators of TLR4 expression and signaling thereby preventing excessive activation of pro-inflammatory cytokines. We hypothesize, that suppression of LPS induced induction of miR-155 and miR-146a, by Morphine and TAT, deregulates the inhibitory feedback loop resulting in sustained TLR4 expression and signaling (Fig. 1). In Specific aim 1: We will test the hypothesis that LPS induction of miR-155 inhibits TLR4 expression and establish that morphine and TAT-mediated attenuation of miR-155 results in greater and sustained TLR4 expression. Specific aim 2: We will test the hypothesis that LPS induction of miR-155 and miR-146a acts as negative feedback regulators of TLR4 signaling and morphine and TAT modulation of miR-155 and 146a results in persistent TLR4 signaling. Specific Aim 3: Determine the mechanism by which Morphine and TAT modulate LPS induced miR-155 and 146a expression. Specific Aim 4: Determine the mechanism and consequence of morphine and TAT modulation of LPS induced miR-155 and miR-146a in vivo using WT and Transgenic animal models. The results from these studies will allow for the development of new therapeutic strategies to attenuate immune activation and reverse HIV disease progression both in HIV infected patients and in HIV infected drug abusing population. PUBLIC HEALTH RELEVANCE: The latest statistics on the world epidemic of AIDS &HIV (UNAIDS/WHO, November 2005) report that at least 40 million people are infected with HIV with nearly 6 million cases of AIDS world-wide. There is a strong correlation between chronic drug use and increase susceptibility to HIV infection. Chronic drug users accounts for approximately a third of all cases of AIDS in the USA and the progression to AIDS dementia is markedly accelerated in opiate drug abusers. This proposal postulates that modulation of microRNAs, mir-155 and miR-146a, by opioid drug abuse and HIV1- TAT may be a plausible mechanism for the persistent immune activation in these patients. Until now there are no published studies implicating miRs in the dysregulated immune activation observed in HIV infected drug abusing population. These studies will allow for the delineation of the mechanisms and allow for the development of new therapeutic strategies to attenuate immune activation and reverse HIV disease progression both in HIV infected patients and in HIV infected drug abusing population.